?-Synuclein (?-Syn) is a protein abundantly distributed in presynaptic terminals. Lewy bodies observed in Alzheimer?s disease (AD), Parkinson?s disease, dementia, and other neurodegenerative conditions contain moderate to high percentages of ?-Syn. Accumulating evidence suggest that ?-Syn accumulation characterizes AD, and a role for neurotoxic ?-Syn species in AD neuropathology has been proposed. Therefore, keeping the right conformation of ?-Syn can contribute to AD prevention. Meanwhile, neuronal soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) are essential for communication between neurons. ?-Syn has been shown to function in promoting the assembly of SNARE complex by clustering synaptic vesicles. This physiological function is highly dependent on binding to the synaptic vesicle membrane. Several studies have suggested that maintaining SNARE function is also important for preventing neurodegenerative diseases including AD. We hypothesize that a membrane environment containing lysophospholipids (lysoPLs) at the presynaptic terminal preserves the right conformation of ?-Syn, allowing it to perform its function in membrane fusion. Here, we propose two Specific Aims to address our hypothesis: (1) Study early transient oligomerization of ?-Syn influenced by interaction with lysoPLs and proteins at the presynaptic terminal; (2) Determine the roles of lysoPLs on ?-Syn?s function in SNARE-mediated membrane fusion. To accomplish these aims, we will use multiple single-molecule assays, including single-particle nanopore detection, single-molecule fluorescent imaging, and single-vesicle fusion. Our single-vesicle fusion assays are ideal tools to reveal the physiological function and to assess the toxicity of various ?-Syn species on synaptic transmission. In addition, the involvement of phospholipids and proteins related to synaptic transmission in our studies of ?-Syn oligomerization or aggregation will be further elucidated. The results of this project will advance our understanding of the normal and pathological function of ?-Syn in the brain and reveal novel therapeutic targets for the treatment of neurodegenerative diseases characterized by abnormal accumulation of ?-Syn, including AD.